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ADCY9 / Adenylate cyclase type 9 · IHC design guide
Design Immunohistochemistry for ADCY9
Plan chromogenic ADCY9 IHC on paraffin sections using high staining in gallbladder glandular cells as a reference (HPA tissue IHC) and 1:50–1:200 as the catalog antibody’s starting dilution (datasheet). Assess the cytoplasmic tissue pattern (HPA tissue IHC), consider the membrane topology (UniProt), and include a no-primary control (datasheet).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
ADCY9 Immunohistochemistry Experimental Design Guide
Expected localisation, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before staining
| Expected localisation | Cytoplasmic in tissue (HPA tissue IHC); membrane protein (UniProt) | |
| Staining pattern | Cytoplasmic staining across most tissues (HPA tissue IHC) | |
| Antigen retrieval | Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen) | |
| Positive control | Gallbladder+4 more · see all | |
| Negative control | Adipose tissue+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A06038-1) | |
| Caveat | Staining has medium concordance with RNA expression (HPA tissue IHC) | |
| Regulation | Expression regulation is not annotated (UniProt) | |
| Isoform / epitope | No annotated isoforms; epitope side matters across 12 transmembrane spans (UniProt) |
Section 1
Recommended ADCY9 IHC & IF Protocols
The catalog antibody’s IHC-P protocol is accompanied by published ADCY9 workflows for colon cancer tissue microarrays and lung adenocarcinoma sections (PMC5759510; PMC10089850).
| Sample | Paraffin-embedded human breast carcinoma tissue; fixative not specified (datasheet A06038-1) |
| Fixation | Image fixative and duration unreported (datasheet A06038-1); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat-induced epitope retrieval in citrate buffer, pH 6.0, 20 min at 95–98 °C (standard rule: cytoplasmic / membrane antigen) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% normal serum of the secondary host, 30 min, room temperature (standard) |
| Primary antibody | Rabbit anti-ADCY9, 1:50-1:200 (datasheet A06038-1) |
| Primary incubation | Overnight at 4 °C (standard) |
| Detection | HRP-polymer secondary, DAB chromogen 5–10 min (standard) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | ADCY9-positive staining in glandular cells of gallbladder (HPA tissue IHC: High). HPA tissue profile: Cytoplasmic expression in most tissues. No signal in the no-primary control. |
💡Decision noteStart with citrate pH 6.0 HIER at 95–98 °C for 20 min for the catalog antibody (page retrieval rule; UniProt O60503: cell membrane).
Section 2
What Is the Expected ADCY9 Staining Pattern?
ADCY9 is a cell membrane protein with 12 transmembrane segments (UniProt O60503 topology). For IHC, expect predominantly cytoplasmic staining across many tissues (HPA: tissue IHC profile), with strong staining in gallbladder glandular cells (HPA: High). HPA rates its tissue IHC evidence Approved, with medium consistency against RNA data and external verification pending (HPA: tissue IHC reliability).
| Strong cytoplasmic staining in gallbladder glandular cells. | This matches the clearest supplied tissue IHC positive (HPA: High in gallbladder glandular cells). Score the glandular cells and their staining intensity, rather than treating any brown deposit in the section as ADCY9. |
| Cytoplasmic staining with some membrane accentuation in the expected cells. | Cytoplasmic staining fits the tissue IHC profile (HPA: cytoplasmic expression in most tissues). Membrane accentuation is biologically plausible (UniProt O60503: cell membrane; HPA: supported plasma membrane location in ICC-IF), but a crisp membrane rim is not required by the tissue IHC record. |
| Predominantly nuclear staining, with little staining in the expected compartment. | Treat this as an unsupported pattern and investigate an artefact or off-target signal: the supplied localization evidence describes cell membrane and cytoplasmic staining, not nuclear localization (UniProt O60503: cell membrane; HPA: tissue IHC profile and subcellular summary). |
| Prominent staining in adipocytes or heart cardiomyocytes. | These specific cell populations are reported as not detected (HPA: adipocytes and cardiomyocytes). Check morphology and detection background before interpreting the signal as ADCY9; staining in an unexpected cell type can reflect cross-reactivity or endogenous detection activity. |
| Diffuse staining across the section, or no staining in gallbladder glandular cells. | Diffuse deposit that obscures cell boundaries is difficult to score; review a reagent control and the chromogenic detection steps (general IHC practice). An absent signal in the supplied high-positive cell population calls for a run-quality check (HPA: High in gallbladder glandular cells). |
💡Expected ADCY9 appearanceCall a section positive when glandular cells show interpretable, predominantly cytoplasmic staining, strongest in a gallbladder positive control (HPA: cytoplasmic tissue IHC profile; High in gallbladder glandular cells); widespread diffuse deposit or dominant nuclear staining is a suspect pattern against the supplied localization evidence (UniProt O60503: cell membrane; HPA: tissue IHC and subcellular profiles).
| Tissue and cell choice | Gallbladder glandular cells offer a high-staining reference; adrenal, appendix, breast, colon and duodenal glandular cells are medium-staining alternatives (HPA: tissue IHC levels). Use the annotated cell population when comparing slides, since intensity can differ by tissue. |
| Membrane topology versus IHC appearance | ADCY9 has 12 transmembrane segments and long cytoplasmic regions (UniProt O60503 topology). Tissue IHC nevertheless reports cytoplasmic expression in most tissues (HPA: tissue IHC profile); do not reject an otherwise matching slide solely because the membrane edge is indistinct. |
| Strength of localization evidence | Plasma membrane localization is supported in ICC-IF, while cytosol localization is uncertain in that assay (HPA: subcellular location). Tissue IHC separately reports a cytoplasmic profile (HPA: tissue IHC). Keep those assay-specific descriptions distinct when interpreting chromogenic sections. |
| Antibody evidence | Two listed antibodies have Approved IHC status, HPA041328 and HPA044225 (HPA: antibody validation). The overall tissue profile has medium consistency with RNA data and awaits external verification (HPA: tissue IHC reliability); use controls when assigning a new cell-level pattern. |
| IF/ICC: what pattern belongs on its own guide page? | Mainly plasma membrane and cytosol staining is reported, with plasma membrane supported and cytosol uncertain; primary cilium and basal body are additional uncertain locations (HPA: subcellular summary). This describes IF/ICC interpretation and supplies no IHC-P protocol option. |
| Situation | Likely cause | Next action |
|---|---|---|
| No signal in gallbladder glandular cells. | A high-positive reference has failed to stain (HPA: High in gallbladder glandular cells); the cause cannot be assigned from the tissue record alone. | Check section integrity, antibody and detection setup, and a concurrent positive control (general IHC practice). Review retrieval conditions as a general workflow check; ADCY9-specific retrieval sensitivity is unreported. |
| Only a weak cytoplasmic signal; no sharp membrane rim. | The observed tissue IHC profile is cytoplasmic, although ADCY9 is a membrane protein (HPA: tissue IHC profile; UniProt O60503: cell membrane). | Assess the expected cell population against control tissue before calling the stain failed (HPA: High in gallbladder glandular cells). Do not require a sharp membrane outline for a positive tissue IHC call. |
| Nuclei dominate the stain. | Dominant nuclear localization is unsupported by the supplied localization records (UniProt O60503: cell membrane; HPA: tissue IHC and subcellular profiles). | Compare a reagent control, inspect morphology, and review primary antibody and detection settings (general IHC practice). Treat the nuclear pattern as suspect until the expected cell-level pattern is demonstrated. |
| Strong staining appears in adipocytes or cardiomyocytes. | Those cell types are listed as not detected, even though UniProt reports ADCY9 detection in heart tissue more broadly (HPA: tissue IHC negatives; UniProt O60503: tissue specificity). | Confirm which cells carry the signal and compare a negative reagent control (general IHC practice). Do not equate tissue-wide detection with a positive cardiomyocyte IHC result. |
| Brown deposit is diffuse or follows section edges. | The deposit may reflect background or endogenous chromogenic activity rather than cell-specific staining (general IHC practice). | Review blocking, washing, detection exposure and the reagent control (general IHC practice). Score ADCY9 only where cellular staining remains interpretable against the expected tissue pattern (HPA: tissue IHC profile). |
| A new tissue shows a different cell pattern. | HPA reports medium consistency between staining and RNA data, with external verification pending (HPA: tissue IHC reliability). The supplied examples do not establish every cell population's pattern. | Record the cell type and compartment, compare supplied positive and negative references, and seek independent confirmation before assigning ADCY9 specificity (HPA: tissue IHC levels and reliability). |
Experiment evidence
IHC & IF images showing ADCY9 staining
A06038-1 · IHCImmunohistochemistry (IHC) analyzes of ADCY9 pAb in paraffin-embedded human breast carcinoma tissue at 1:50,showing cytoplasm and membrane s
A06038-1 · IHCImmunohistochemistry (IHC) analyzes of ADCY9 pAb in paraffin-embedded human stomach carcinoma tissue at 1:50,showing cytoplasm and membrane
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000162104-ADCY9/tissue/cerebral+cortex#imid_10697743
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000162104-ADCY9/tissue/colon#imid_10697669
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000162104-ADCY9/tissue/liver#imid_10697678
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000162104-ADCY9/tissue/kidney#imid_10697687
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA041328; green = ADCY9, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA041328; green = ADCY9, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for ADCY9 IHC & IF
🧪Run gallbladder first: glandular cells should stain (HPA: High in gallbladder glandular cells). Use adipose tissue as a negative comparator (HPA: Not detected in adipocytes); no internal ADCY9-negative cell type is established for gallbladder, so any candidate internal negative cells must be independently validated and should show no specific staining (HPA: gallbladder row; standard IHC practice).
Positive control tissue: Gallbladder (Glandular cells, HPA High)
Negative control tissue: Adipose tissue (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show ADCY9 in A-431, U-251MG, U2OS, ASC52telo, RPTEC/TERT1, hTERT-RPE1 (serum starved), KOLF2.1J, with annotated localisation: Plasma membrane (supported), Cytosol (uncertain) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and concentration-matched nonimmune rabbit IgG as the isotype control (selected IHC caption: rabbit pAb with anti-rabbit secondary; standard IHC practice); use ADCY9 knockout tissue or a validated immunizing-peptide block as a biological negative (standard IHC practice). In gallbladder, check endogenous peroxidase and biotin background before interpreting avidin-peroxidase signal (selected IHC caption: avidin-peroxidase detection; standard IHC practice).
⚠️Feasibility: A target-specific fixation window and antigen-retrieval dependence are unreported in the supplied evidence; the selected A06038-1 paraffin-section caption does not report a fixative (selected IHC caption). The evidence does not establish whether frozen sections or IF/ICC are easier; supported plasma-membrane localization can guide IF interpretation, while cytosolic and ciliary localization is uncertain (HPA subcellular). In gallbladder, inspect luminal material and detection background before scoring glandular staining (HPA: High in gallbladder glandular cells; standard IHC practice).
HPA tissue IHC evidence for ADCY9
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Approved — Medium consistency between antibody staining and RNA expression data. Pending external verification.). Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Gallbladder | Glandular cells | High | Protein (IHC) | HPA → |
| Adrenal gland | Glandular cells | Medium | Protein (IHC) | HPA → |
| Appendix | Glandular cells | Medium | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | Medium | Protein (IHC) | HPA → |
| Breast | Glandular cells | Medium | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adipose tissue | Adipocytes | Not detected | Protein (IHC) | HPA → |
| Heart muscle | Cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Ovary | Ovarian stroma cells | Not detected | Protein (IHC) | HPA → |
| Prostate | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Smooth muscle | Smooth muscle cells | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced ADCY9 IHC Tips
Troubleshoot ADCY9 staining in paraffin sections by checking retrieval, cellular pattern and controls before assigning biological meaning.
What retrieval should I try when ADCY9 staining is weak in paraffin sections?
Use citrate buffer at pH 6.0 for heat retrieval at 95–98 °C for 20 min (page retrieval rule: cytoplasmic/membrane antigen). Allow sections to cool in buffer, then compare a matched positive control and a no-primary control under identical detection conditions (standard IHC practice). The selected A06038-1 image shows staining in paraffin-embedded breast carcinoma at 1:50, but its caption does not specify retrieval (A06038-1 caption). If staining remains weak, test a higher-pH retrieval buffer on adjacent sections while keeping antibody concentration and detection constant (standard IHC practice). Reject conditions that raise diffuse staining or damage tissue architecture (standard IHC practice).
Could fixation explain inconsistent ADCY9 staining between paraffin blocks?
Target-specific fixation sensitivity is unknown because the supplied ADCY9 tissue-IHC caption does not state a fixative (A06038-1 caption). Record the fixative and fixation duration for each block, and compare sections with matched processing histories before changing the staining method (standard IHC practice). Inconsistent fixation can alter antigen access and tissue morphology in paraffin IHC, so assess both staining intensity and structural preservation (standard IHC practice). Run the same retrieval, 1:50 starting dilution, and detection sequence across comparable sections when evaluating A06038-1 (A06038-1 caption; standard IHC practice). Do not infer an ADCY9-specific fixation effect from tissue-expression patterns or protein topology (HPA tissue IHC; UniProt O60503 topology).
Is cytoplasmic ADCY9 staining plausible for a membrane protein?
ADCY9 is annotated at the cell membrane and has 12 transmembrane segments with large cytoplasmic regions (UniProt O60503 topology). Its selected paraffin-section image describes cytoplasmic and membrane staining in breast carcinoma, while HPA reports cytoplasmic staining in most tissues (A06038-1 caption; HPA tissue IHC). Assess whether chromogen follows cell borders, cytoplasm, or both, and compare that pattern with a no-primary section (standard IHC practice). A wholly nuclear pattern would conflict with the supplied localisation evidence and warrants a specificity check before scoring (UniProt O60503 localisation; HPA subcellular). Preserve compartment-specific observations rather than combining all brown signal into one positive category (standard IHC practice).
How should unknown antibody epitope placement affect ADCY9 IHC optimisation?
The supplied record lists 0 annotated isoforms and does not identify the catalog antibody’s epitope (UniProt O60503 record; A06038-1 caption). ADCY9 spans the membrane 12 times and contains cytoplasmic and extracellular segments, so epitope accessibility cannot be assigned without antibody-specific mapping (UniProt O60503 topology). Keep the recommended citrate pH 6.0 retrieval as the starting condition, then compare adjacent sections if signal remains weak (page retrieval rule; standard IHC practice). Glycosylation is annotated at residues 206, 955, and 964, but these annotations alone do not establish interference with this antibody (UniProt O60503 glycosylation). Interpret retrieval changes against morphology and no-primary staining (standard IHC practice).
How can I check ADCY9 localisation by multiplex IF without misreading background?
Use IF as a separate validation experiment; the selected A06038-1 evidence describes paraffin-section chromogenic IHC, not IF performance (A06038-1 caption). Multiplex ADCY9 with a marker for the expected cell population, such as a smooth-muscle marker when examining airway smooth muscle, where expression was reported highest in lung (UniProt O60503 tissue specificity; standard IF practice). Choose spectrally separated fluorophores, favouring a far-red ADCY9 channel if the specimen has strong short-wavelength autofluorescence (standard IF practice). If the antibody epitope is cytoplasmic, use controlled permeabilisation; if extracellular, first assess staining without permeabilisation because epitope placement is unspecified (UniProt O60503 topology; standard IF practice). Include single-stain and no-primary controls (standard IF practice).
What should I change when ADCY9 chromogen appears throughout the section?
First compare the stained section with a no-primary control; the selected A06038-1 image used PBS in place of primary antibody for its negative control (A06038-1 caption). Apply a peroxidase block before avidin–peroxidase detection and assess whether residual endogenous enzyme contributes to diffuse chromogen (A06038-1 caption; standard IHC practice). Reduce primary antibody concentration from the caption’s 1:50 condition in a controlled dilution series if background rises with antibody exposure (A06038-1 caption; standard IHC practice). Check blocking, wash quality, section drying, and chromogen development on adjacent sections while holding retrieval constant (standard IHC practice). Score only cellular staining distinguishable from the matched control (standard IHC practice).
How should I quantify ADCY9 staining across heterogeneous tissue sections? ⚠ ANSWER MARKED FOR VERIFICATION
Define the cell population and compartment before scoring, because the selected paraffin image reports both cytoplasmic and membrane staining (A06038-1 caption). For cellular expression, record the percentage of positive cells by intensity category and calculate an H-score, or report percentage positive with the scoring threshold stated (standard IHC practice). If counting spatially distributed positive cells, report density per mm² of viable tissue and exclude empty spaces and damaged areas (standard IHC practice). Normalise comparisons to the same cell type or viable tissue area, with identical exposure to retrieval and chromogen development (standard IHC practice). Keep membrane and cytoplasmic scores separate when their patterns differ (standard IHC practice).
How do I distinguish credible ADCY9 staining from section artefacts?
Credible staining should have a reproducible cellular pattern compatible with membrane or cytoplasmic localisation and exceed the matched no-primary control (UniProt O60503 localisation; A06038-1 caption; standard IHC practice). Check the relevant cell population: HPA reports high staining in gallbladder glandular cells and no detection in adipocytes, although its tissue-IHC reliability is Approved with pending external verification (HPA tissue IHC). Treat isolated nuclear staining, section-edge concentration, and signal over necrosis as suspect until controls and repeat sections support them (UniProt O60503 localisation; standard IHC practice). Check endogenous peroxidase contribution when using chromogenic enzyme detection, especially if the no-primary section develops signal (A06038-1 caption; standard IHC practice). Base conclusions on morphology, controls, and replicated cell-specific staining (standard IHC practice).
Boster reagents
Best ADCY9 / Adenylate cyclase type 9 IHC Antibodies
A06038-1 has pictured ADCY9 IHC staining in paraffin-embedded human breast and stomach carcinoma (IHC captions). The catalog lists human, mouse and rat reactivity; no IF image is supplied (catalog reactivity; IF image field).
Real IHC data
Anti-Adenylate cyclase type 9 ADCY9 Antibody
Cat # A06038-1
A06038-1 will render with its human breast carcinoma IHC figure, showing cytoplasmic and membrane staining at 1:50 in a paraffin-embedded section (A06038-1 figure caption). A second caption documents human stomach carcinoma IHC; the catalog lists IHC and WB applications and human, mouse and rat reactivity (A06038-1 IHC caption; catalog applications and reactivity).
Which to pick: For tissue IHC, choose A06038-1, described as a rabbit pAb, for its pictured paraffin-embedded human tissue staining; the fixative is unreported (catalog host; A06038-1 IHC caption). No SKU here has documented IF/ICC validation: A06038-1 lists IHC and WB, with no IF image or dilution (catalog applications; IF fields). For mouse or rat IHC, A06038-1 is the listed option by reactivity, although its pictured IHC evidence is from human tissue (catalog reactivity; A06038-1 IHC captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry O60503 (ADCY9_HUMAN, Adenylate cyclase type 9).
- Human Protein Atlas. ADCY9 tissue IHC expression (reliability: Approved).
- Human Protein Atlas. ADCY9 subcellular location (ICC-IF): Mainly localized to the plasma membrane and cytosol. In addition localized to the primary cilium and basal body..
- Human Protein Atlas. ADCY9 antibody validation summary (2 antibodies).
- Elevated Adenylyl Cyclase 9 Expression Is a Potential Prognostic Biomarker for Patients with Colon Cancer. Medical science monitor : international medical journal of experimental and clinical research 2018 — PMC5759510.
- ADCY9 functions as a novel cancer suppressor gene in lung adenocarcinoma. Journal of thoracic disease 2023 — PMC10089850.
- PubMed PMID:9628827 — UniProt-cited evidence.
- PubMed PMID:12972952 — UniProt-cited evidence.
- PubMed PMID:15879435 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for ADCY9 (Adenylate cyclase type 9). Expected localisation Cytoplasmic in tissue (HPA tissue IHC); membrane protein (UniProt). Antigen retrieval: Citrate pH 6.0 HIER, 95–98 °C, 20 min (rule: cytoplasmic / membrane antigen). Validated IHC antibody: A06038-1 at 1:50-1:200 (the product's own IHC validation image is shown). Controls (HPA): positive Gallbladder (Glandular cells); negative Adipose tissue. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human breast carcinoma tissue; fixative not specified (datasheet A06038-1); 2. Fixation: Image fixative and duration unreported (datasheet A06038-1); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat-induced epitope retrieval in citrate buffer, pH 6.0 , 20 min at 95–98 °C…; 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% normal serum of the secondary host, 30 min , room temperature (standard); 8. Primary antibody: Rabbit anti-ADCY9, 1:50-1:200 (datasheet A06038-1); 9. Primary incubation: Overnight at 4 °C (standard). Watch for: Fixation: Matched tissue-IHC evidence does not establish the fixation claim. Validate the specimen-specific method before use. (selected-SKU IHC image A06038-1); Caveat: Staining has medium concordance with RNA expression (HPA tissue IHC); Isoform / epitope: No annotated isoforms; epitope side matters across 12 transmembrane spans (UniProt); Regulation: Expression regulation is not annotated (UniProt). Evidence: UniProt O60503, HPA tissue IHC · Approved, HPA ICC-IF · Mainly localized to the plasma membrane and cytosol. In addition…, Datasheet A06038-1, PMC5759510, PMC10089850. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/ADCY9/immunohistochemistry-ihc-guide.